Neuroticism, Acculturation and the Cortisol AwakeningResponse in Mexican American Adults

Deborah Mangold, Ph.D.*,1, Jim Mintz, Ph.D.2, Martin Javors, Ph.D.2, and Elise Marino, B.A.1

1The University of Texas at San Antonio, San Antonio, Texas 782492The University of Texas Health Science Center at San Antonio, San Antonio, Texas

AbstractNeuroticism is associated with greater susceptibility to the adverse effects of stress and greaterexposure to the stressors associated with acculturation in U.S. born Mexican Americans.Neuroticism and acculturation have been associated with injury to crucial stress response systemsand are known risk factors for certain mood and anxiety disorders. The purpose of the currentstudy was to examine the effects of neuroticism, and acculturation on the cortisol awakeningresponse (CAR) in healthy Mexican-American adults. Salivary cortisol samples were collected atawakening and 30, 45, and 60 minutes thereafter, on two consecutive weekdays from 59 healthyMexican American adult males (26) and females (33), ages 18 to 38 years. Participants wereassessed for level of neuroticism and acculturation. Data were analyzed using a mixed effectsregression model with repeated measures at four time points. Results showed a significantNeuroticism × Acculturation × Time interaction. The CAR was virtually eliminated in highlyacculturated Mexican Americans with greater Anglo orientation and high neuroticism comparedwith less acculturated Mexican Americans with greater Mexican orientation and lowerneuroticism. Findings suggest that some Mexican Americans with high levels of neuroticism maybe particularly susceptible to certain challenges and stressors associated with acculturation leadingover time to the development of allostatic load, desensitization of the Hypothalamic CRF systemand attenuation of the CAR.

KeywordsNeuroticism; Acculturative Stress; Acculturation; HPA axis; Cortisol; Mexican-Americans

IntroductionNeuroticism is one of five independent personality dimensions that are heritable and stableover the adult life span (Costa and McCrae, 1992). Neuroticism is defined as the pronenessof the individual to experience negative affective states, and is associated with increasedexposure to stressful life events (Bolger and Zuckerman, 1995; Felsten, 2004, 2002) greatersusceptibility to the adverse effects of stress (Kendler et al., 2004; Ormel et al., 2001) and

© 2011 Elsevier Inc. All rights reserved.*Corresponding Author Deborah L. Mangold, Ph.D., Associate Professor of Psychology, The University of Texas at San Antonio, HSS4.04.21, 6900 North Loop 1604 West, San Antonio, Texas 78249, Tele: (210) 458-7405, Fax: (210) 458-5728, dmangold@utsa.edu.Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to ourcustomers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review ofthe resulting proof before it is published in its final citable form. Please note that during the production process errors may bediscovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

NIH Public AccessAuthor ManuscriptHorm Behav. Author manuscript; available in PMC 2013 January 1.

Published in final edited form as:Horm Behav. 2012 January ; 61(1): 23–30. doi:10.1016/j.yhbeh.2011.09.009.

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thus, a major risk factor for mood and anxiety disorders (DeGraaf et al., 2002; Fanous et al.,2002; Roberts and Kendler, 1999).

The relationship between neuroticism and stress may be of particular importance in MexicanAmericans given that studies suggest a number of Hispanics face unique stressors associatedwith some aspects of acculturation (Finch et al., 2000; Huebner et al., 2005; Mena et al.,1987; Miranda and Matheny, 2000). Acculturation involves adaptation into a host culture(Mena et al., 1987) and acculturative stress includes the psychological, somatic, and socialstressors that are associated with the process of acculturation (Bernal and Santiago, 2006;Berry, 1980; Cervantes and Castro, 1985). Neuroticism is associated with more exposure toacculturative stress (Mangold et al., 2007) and greater exposure to acculturative stress isassociated with increased risk for mood and anxiety disorders in U.S. born MexicanAmericans (Finch, et al., 2000; Matheson et al., 2008; Romero and Roberts, 2003; Thomanand Suris, 2004). Indeed, exposure to acculturative stress is more likely to predict depressivesymptomatology in Mexican Americans with high neuroticism (Hovey and King, 1996;Huebner et al., 2005; Salgado de Snyder, 1987). These findings raise the possibility thatneuroticism may be a risk factor that significantly increases susceptibility to the adverseeffects of stressors associated with acculturation.

There is growing interest in the extent to which chronic exposure to stressful life eventsleads to dysregulation of crucial biological stress response systems associated with increasedvulnerability for mood and anxiety disorders. Stress-induced dysregulation of theHypothalamic-Pituitary-Adrenal (HPA) axis is one neurobiological pathway that may linkneuroticism with increased risk for mood and anxiety disorders in Mexican Americansexposed to high levels of acculturative stress. The cortisol awakening response (CAR) hasreceived increasing attention as a useful method to assess the integrity of the HPA axis. TheCAR is a reliable biological marker of HPA activity, dependent on a moderate geneticinfluence (Bartels et al., 2003; Schmidt-Reinwald et al., 1999; Wust et al., 2000a) andchanges in the CAR can yield important information regarding the relationship betweenaltered stress responsivity and attenuation of the awakening portion of the cortisol circadianrhythm. The sensitivity/capacity of the adrenal cortex is proposed to play a crucial role inthe magnitude of the CAR (Kudielka and Kirschbaum, 2003; Pruessner et al., 1997, 1999).Thus, examinations of the extent to which neuroticism may link high levels of acculturativestress with dysregulations of the CAR and greater risk for mood and anxiety disorders inMexican Americans, may be of particular interest.

While studies examining the effects of neuroticism on the CAR in Mexican Americans arenon-existent, results from studies in non-Hispanics are mixed showing an enhanced CAR(Polk et al 2005; Portella et al, 2005; Vedhara et al., 2006), diminished CAR (Hauner et al.,2008), and no association between neuroticism and the CAR (Chan et al., 2007; van Santenet al., 2010). Inconsistent results may be due to differences in cortisol measurement and thediagnostic status of the samples examined (i.e., subjects with and without a formal diagnosisof depression versus subclinical symptomatology). Moreover, no available studies haveexamined the effects of neuroticism on the CAR while controlling for childhood trauma, afactor known to alter the HPA axis and the CAR (Heim and Nemeroff, 2001; Stetler andMiller, 2005; Mangold et al., 2010).

Equally lacking are studies examining the effects of unique forms of cultural stress on theCAR. Studies conducted in our laboratory showed a positive association betweenacculturative stress and neuroticism in Mexican Americans (Mangold et al., 2007 ) and thatgreater acculturative stress and more Anglo-orientation is associated with attenuation of theCAR, after controlling for the effects of childhood trauma and independent of a formaldiagnosis of depression (Mangold et al., 2010). These findings suggest that similar to

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childhood trauma (Mangold et al., 2010), and subclinical symptomatology (Mangold, et al.,2011; Dedovic et al., 2010), acculturative stress is associated with attenuation of the CAR inadults. This perhaps is not surprising given that theorists propose greater acculturation andmore Anglo orientation may contribute to the erosion of crucial protective factors in someindividuals, resulting in greater susceptibility to the adverse effects of chronic exposure toacculturative stress (Finch and Vega, 2003; Vega and Amaro, 1994). The absence ofprotective factors, in turn, may lead to the development of allostatic load, decreased adrenalcapacity over time (McEwen and Lasley, 2003) and attenuation of the CAR. Indeed,theorists have proposed the “attenuation hypothesis” as one possible explanation that isconsistent with the differences observed in the CAR in these studies. The “attenuationhypothesis” suggests that following chronic exposure to stress the hypothalamic CRF systemmay undergo desensitization, transitioning over time from a system with increased stressresponsivity to a system with attenuated cortisol responses to stress and an attenuated CAR(Gunnar and Vazquez, 2001; Heim et al., 2008; Susman, 2006; Trickett et al., 2010). Resultsfrom a meta-analysis showing a negative correlation between length of time following theonset of chronic stress and the magnitude of the CAR provide additional support for thiscontention (Miller et al., 2007).

To date, there is a notable absence of studies examining the effects of neuroticism on theCAR in Mexican Americans and the majority of previous studies have failed to examine theeffects of neuroticism while adequately controlling for factors known to alter the CAR (e.g.,depression and childhood trauma, cortisol measurement with respect to time of awakening).Moreover, studies designed to distinguish the effects of neuroticism from the effects ofacculturative stress on the CAR in Mexican Americans may be of particular importance. Inview of the fact that neuroticism is associated with increased reactivity and susceptibility tothe negative effects of psychosocial stressors, it is plausible that Mexican Americans withhigh levels of neuroticism, who are exposed to chronic acculturative stress, may represent anextraordinarily vulnerable group at high risk for attenuation of the CAR. Therefore, the aimsof the current investigation were to: (1) examine the effects of neuroticism on the CAR inMexican Americans utilizing a carefully constructed sample monitoring system; (2) examinethe effects of neuroticism on the CAR while carefully screening for a diagnosis ofdepression and controlling for the effects of childhood trauma, age and sex and; (3)distinguish the effects of neuroticism from the effects of acculturative stress on the CAR.Based on previous findings supporting the “attenuation hypothesis”, we hypothesized asignificant time × neuroticism × acculturation interaction where neuroticism would beassociated with a significantly attenuated CAR in highly acculturated Mexican Americanswith greater Anglo orientation after controlling for childhood trauma, depression, age andsex.

Materials and MethodsParticipants and Study Design

The study was approved by the University of Texas Institutional Review Board, and allparticipants gave written, informed consent prior to participation. Participants of Mexicandescent (n=59), aged 18 to 38, were recruited from the San Antonio metropolitan area,through advertisements in the community and local college campuses. Specific details ofprocedures for the current study have been reported elsewhere (Mangold et al., 2010).During an initial visit to the laboratory participants underwent a screening interview and abattery of self-report assessments designed to identify and exclude factors known topotentially affect the HPA axis including: lifetime depression (Bhagwager et al., 2005; Sheaet al., 2007); use of oral contraceptives in the past 60 days (Meulenberg and Hofman, 1990;Pruessner et al., 1997, 1999); current pregnancy (Meulenberg and Hofman, 1990); menstrualcycle abnormalities (Bao et al., 2003, 2004; Suh et al., 1988); strenuous aerobic exercise

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(Hansen et al., 2008; Kanaley et al., 2001; Kelly et al., 2008); major medical conditions orhistory of head trauma; use of medications; severe obesity and current alcohol or other druguse disorders (Hansen et al., 2008; Huizink et al., 2006; Wand and Dobs, 1991). In addition,participants reporting abnormal sleeping patterns (Lasikiewicz et al., 2008), shift work orovertime (Lundberg and Hellstrom, 2002) were excluded from participation in the study(Clow et al., 2004; Hanrahan et al., 2006).

Psychometric AssessmentPersonality/Neuroticism—Personality was measured with the Revised NEO PersonalityInventory (NEO PI-R; Costa and McCrae, 1992) a 240-item self-report measure of the fivefactors of personality including Neuroticism, Extraversion, Openness to Experience,Agreeableness and Conscientiousness (Costa et al., 1991; Costa and McCrae, 1992a, b;McCrae and Costa, 1983). Reliability for the current sample was strong (α =.92) andconsistent with reliability reported in a previous study conducted in our laboratory,employing the NEO with healthy, English-speaking Mexican and Mexican American youngadults (Mangold et al., 2007).

Acculturation—Acculturation was measured with the revised Acculturation Rating Scalefor Mexican Americans (ARSMA-II; Cuéllar et al., 1995). The ARSMA-II measuresacculturation by assessing scores on two subscales: The Anglo Orientation Scale (AOS) andthe Mexican Orientation Scale (MOS). The ARSMA-II has strong psychometric propertieswith good reliability for the current sample α =.89).

Depression—Participants were screened for depression using the Hamilton DepressionInventory Short Form (HDI-SF; Reynolds and Kobak, 1995a) and excluded fromparticipation based on a score of 10 or greater as recommended by Reynolds and Kobak(1995b) indicating a strong likelihood of mild, moderate or severe depression. Reliability forthe current sample was α=.78.

Substance Abuse/Dependence—Participants were screened and excluded for alcoholand other drug use disorders using the World Health Organization Alcohol, Smoking andSubstance Involvement Screening Test (WHO ASSIST; Humeniuk et al., 2008). TheASSIST assesses the frequency of lifetime and recent use (within the past 3 months) andproblems associated with the use of alcohol and other drugs of abuse (e.g., cannabis,cocaine, stimulants, etc.). Reliability for the current sample was α=.80.

Childhood Trauma—The current study assessed exposure to severe childhood stress priorto the age of 16, using the Early Trauma Inventory–Short Form adapted from the clinicianadministered Early Trauma Inventory (ETISR-SF; Bremner et al., 2007). The ETISR-SF hasstrong psychometric properties (Bremner et al., 2007; Hyman et al., 2005) with goodreliability for the current sample (α = .82).

Ethnicity/Nativity—Participants reporting an Hispanic origin other than Mexican orMexican-American were excluded from the study. Participants were determined to be ofMexican descent if they reported that both biological parents and both maternal and paternalgrandparents were of Mexican descent.

Measurement of the CAR—Participants were free to wake up as usual (using an alarmor spontaneously), and instructed to start sampling immediately at awakening (T0), and T30,T45 and T60 minutes following awakening. Sampling was completed on two consecutiveweekdays. Wake-up times were logged along with sampling times for comparison withelectronic monitoring data. All participants completed the cortisol protocol within 3 weeks

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of screening. To avoid contamination of cortisol samples, participants were instructed torefrain from brushing their teeth, eating/drinking, strenuous exercise, alcohol, smoking andcaffeine during sampling (Badrick et al., 2007; Clow et al., 2004). Participants placed a rollof cotton in their mouths, chewing on it until it became saturated, before returning to thesalivette (Sarstedt; Rommelsdorft, Germany). Participants were instructed to remain sittingupright in bed until the second saliva sample was obtained, when they were free to followtheir normal weekday routine during the sampling period. Cortisol was assessed in femalesduring the early follicular phase of their menstrual cycle (determined by menstrual diary)employing a conservative approach to control for possible effects of menstrual cycle phaseon the CAR (Bao et al., 2003, 2004).

We utilized electronic monitoring devices together with self-reported time of sampling todetermine concordance between monitored times and self-reported times, detect deviationsfrom the protocol and maximize accuracy in the documentation of sample times (Jacobs etal., 2005; Mangold et al., 2010, 2011). Participants collected saliva samples according toidentical protocols on two separate, consecutive days. Salivettes were stored in a vialequipped with an electronic medication event monitoring system (MEMS; Aardex; Zug,Switzerland) as recommended for use in an ambulatory setting by (Kudielka et al., 2003).The MEMS is designed to compile the dosing/sampling-compliance histories of participants.This system includes a standard plastic vial with cap that contains a threaded opening and aclosure that contains a micro-electronic circuit that registers times when the closure isopened and when it is closed. Time-stamped medication events are stored in the unit forlater software analyses to maximize accuracy of recorded awakening/sampling timesincluded in later analyses.

Hormonal Assays—Participants stored the samples in their home freezers until they werereturned to the laboratory the next day and stored at −80 °C until analyzed, when they werethawed at room temperature and then centrifuged at 2500 g for 15 minutes at −4°C. Salivarycortisol levels were assayed in duplicate using high sensitivity enzyme immunoassays(Diagnostic Systems Laboratories; Webster, Texas) with a mean lower sensitivity limit of0.11 µg/dL, standard curve range from 0.1 to 10µg/dL. The intra-assay and inter-assaycoefficients of variation were less than 5% at all levels of the calibrator curve. Theconcentration of cortisol in saliva was expressed as µg/dL. To minimize the potential effectsof exposure to stressful events during the sampling period participants who were currentlystudents were not sampled the week prior to scheduled class examinations. Participants wereasked to record stress exposures/daily hassles (Van Eck et al., 1996), sleep disturbances(Lasikiewicz et al., 2008) and protocol noncompliance (teeth brushing, eating, etc) known toaffect HPA axis activity during the sampling period. However, there were no problemsrecorded during the sampling period and thus; analyses of data were performed on a finalsample of 59 subjects.

Statistical Analyses—Details of the sample and the analytic approach were presented ina prior publication based on the same sample (Mangold et al., 2010). A total of 65participants were screened for enrollment in the study, and six participants were excludedbased on use of oral contraceptives, prescription medication use, and a diagnosis of majordepression. The analyses are based on data for 59 participants. Descriptive statistics arereprinted here to characterize the sample. Cortisol data were log-transformed to correctpositive skew. The primary statistical design was a mixed effects regression model withrepeated measures at four time points (awakening and 30, 45, and 60 minutes thereafter).Fixed design effects were self-reported Neuroticism (NEO-PI-R; Costa and McCrae, 1992),used as a continuous, dimensional variable (raw scores ranged from 31 to146 and T scoresranged from 22.82 to 85.58), Acculturation (ARSMA-II; Cuéllar et al., 1995), a continuousvariable used in our prior study, and Time (a fixed classification factor with four levels), and

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their interaction. We used a fully crossed design (Neuroticism × Acculturation × Time),focusing on interactions with Time which test effects on the changes in cortisol over time.Continuous variables were centered by subtracting the mean to reduce confounding of lowerand higher order effects (Kraemer and Blasey, 2004).

Modeling time as a fixed classification factor with 4 levels simplified specification of thecovariance structure and presentation of the results, and was supported by the observation ofrelatively little intra-individual variability in the timing of cortisol sampling: 89% of thesamples were obtained within five minutes of the scheduled time, and 95% within 10minutes of protocol. We specified the Kenward-Rogers adjustment for degrees of freedomand a spatial power structure for the covariance matrix with measures at 0, 30, 45 and 60minutes. This is a generalization of first-degree autoregressive structure to the case ofunequally spaced measures. It was preferred to autoregressive, compound symmetry, or anunstructured matrix by the BIC likelihood-criterion provided by SAS MIXED (Littell, et al.,1996, pp. 126–130).

All but three participants provided cortisol data following the same measurement protocolon two consecutive weekdays. We used the average of the day one and day two data becauseaveraging over a period from two to six days increases trait specificity (Clow et al., 2004;Hellhammer et al., 2007; Kunz-Ebrecht et al., 2004, Scholtz et al., 2004; Thorn et al., 2006,2009) and considerably simplifies the covariance structure and analysis. We had noexpectation of meaningful day-to-day variability, and confirmed this by performingpreliminary mixed effects analyses including Day as an additional design factor. These didnot reveal any significant effects involving Day or meaningfully change the results, and arenot reported further. Statistical analyses were done using the SAS 9.2 statistical library.Tests were at unadjusted two-tailed p =.05. To aid interpretation of the Neuroticism byAcculturation interaction, we graphed means at each time point for three patterns of thedimensions: one SD below the mean, at the mean, and one SD above the mean on bothscales.

ResultsParticipant Characteristics

Table 1 presents demographic characteristics of the sample. Participants were single (n =59), healthy, adult males (44 percent) and females (56 percent). Most were first or secondyear college students. They ranged in age from 18–38 years (median = 20 years, M = 22.0,SD = 5.4) and Body Mass Index (BMI) within normal parameters (M = 24.4, SD = 4.2). Alittle over half the sample reported an annual family income of $40,000 or less, and less thanone-fifth reported a household income at or above $80,000. Approximately one-third of thesample were first generation Mexican Americans (born in Mexico and immigrated as a childor as an adult to the United States), approximately one-fourth of the sample were secondgeneration (born in the U.S. of Mexican born parents, and a little over forty percent werethird generation or more (parents and self were born in the United States). The majoritywere bilingual. Participants were generally healthy, with mean scores on the General healthsubscale of the RAND-36 in line with those previously reported for healthy, young adults(M = 77.6, SD = 20.1; Vander Zee et al., 1996). No current or lifetime psychiatric diagnosesor current use of psychotropic medication was reported. Individuals with HDI scores higherthan 10, indicating depressive symptomatology, were excluded from the study, andtherefore, the sample HDI mean (M =2.8, SD =2.4) is lower than normative means reportedfor college-aged participants (M = 5.16, SD = 4.48), reported by Reynolds and Kobak(1995b) and Vander Zee et al. (1996). However, scores on the HDI ranged from 0 to 9.50,suggesting subjects endorsed a range of depressive symptoms.

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Exposure to Early TraumaParticipants, on average, reported exposure to more than five different trauma items acrossall trauma dimensions (M = 5.32, SD = 4.25). This is consistent with epidemiologicalevidence suggesting that over 56% of Americans experience a lifetime trauma, and risk ofexposure to general or complex traumas are reported to be as high as 72% among Latino/Hispanic children (NCTSN; 2005).

NeuroticismTable 2 reports mean raw scores for the NEO PI-R personality factor of Neuroticism andneuroticism facets for the Mexican-American male and female subjects in the currentsample. To our knowledge, normed scores for Mexican-American samples are not yetavailable. Overall raw scores for the current sample were consistent with norms publishedfor a predominately Caucasian sample (Costa and McCrae, 1992a) and with scorespreviously reported for a sample of Mexican American college students (Mangold et al.,2007). There were no significant differences in the neuroticism factor and facets scoresamong males and females.

Characterization of the Cortisol Awakening ResponseSalivary cortisol response for the subject sample was characterized with respect to overallcortisol secretory activity measured in the period 1hr after awakening and the change inconcentration from awakening to 30 min post-awakening (Clow et al., 2004). Calculation ofthe mean changes in cortisol concentration from awakening to 30 min post-awakening fortrial one (12.4 ± 16.5 nmol/l) and trial two (9.9 ± 16.1 nmol/l) compare to what is previouslyreported (9.3 ± 3.1 nmol/l), although with more variation (Clow et al., 2004). Thecorrelation between delta cortisol for days one and two was r = 0.47 (df = 54, p = .0003;Spearman rho = 0.34, p=.01). The simple correlation across all data points (i.e., using 4values per person with N = 59) was r = 0.54 for raw cortisol, and r = 0.42 for log cortisol.

Neuroticism, Acculturation and Cortisol Awakening ResponseTable 3 presents the mixed effects regression analysis. The two factor Acculturation byTime interaction was significant (p = 0.0014), although the Neuroticism by Time interactionwas not. The three-factor Neuroticism by Acculturation by Time interaction was highlysignificant (p = 0.0026) when the Neuroticism and Acculturation scales were included asdimensional covariates. Moreover, when the three subjects who did not provide cortisol dataon Day 2 were omitted from the analyses, the three-factor interaction remained significant(F=4.99, df=3, 154, p=0.0025).

To display the result, we split both the Neuroticism and Acculturation scales at their meansto create dichotomous indicators (low, high), and reanalyzed the data using theseclassifications in a 2 (Neuroticism) × 2 (Acculturation) × 4 (Time) mixed effects regressionmodel. Figure 1 depicts the estimated CAR in the four groups created by crossing thesedichotomous classifications. Participants who were low on both dimensions had a greaterCAR in comparison to the other groups (a test of change over time in this group yielded (F =14.75, df = 3,162, p<0.0001). Those high on one of the two dimensions but low on the otherhad a somewhat attenuated CAR. Their CAR profiles were quite similar, and both indicatedsignificant change over time (F = 10.46 and 8.58 respectively, df = 3, 162, p<0.0001). Incontrast, those with high levels of both Neuroticism and Acculturation showed almost noCAR at all (test for change over time yielded F =1.07, df = 3, 162, p = 0.36). The combinedeffect of being high on both dimensions was particularly evident in the virtual lack ofchange in cortisol seen in this group at 30 minutes after awakening.

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Simple effects analyses indicate a non-significant group difference at Time 0 (F=0.31,p=0.82), and significant differences at 30 (F=3.98, p=0.010), 45 (F=5.51, p=0.0015) and 60minutes (F=3.21, p=0.0263, all numerator df=3). Examination of effect size using Cohen’sd, calculated for the data displayed in Figure 1, suggest differences between the LowAcculturation-Low Neuroticism group (most robust CAR) and the High Acculturation-HighNeuroticism group (most attenuated CAR) are very large (large effect size) at all three timepoints (d=1.24, 1.30, 1.10). Differences between the Low Acculturation-Low Neuroticismgroup and the intermediate groups with mixed features (high on one dimension only) aremoderate (medium effect size) only by 60 minutes (d=0.49, 0.57). Differences betweenthose intermediate groups and the High Acculturation – High Neuroticism group are large,initially at 30 minutes (d=0.96, 1.17).

Finally, we evaluated the robustness of these results when adding age, sex, level of familyincome and a measure of early trauma to the analysis as covariates. The three-factorinteraction of Neuroticism by Acculturation by Time remained significant (F = 4.82, df = 3,159, p =.0031) and the pattern of estimated means was virtually unchanged (the meandifference in estimated means was zero). Moreover, although generational status was highlycorrelated with the ARSMA (r=0.605, df=57, p<.0001) in our sample, when we addedGeneration to the regression model both as a main effect and interaction with time (tocapture effects of Generation on the CAR), Generation had no effect on the three factorinteraction, which was slightly more significant with these two Generation effects in themodel (F=5.12, df=3, 160, p=.0021). Generation itself was not a significant predictor in thismodel (main effect F=0.02, df=1, 57.8, p=0.91, interaction with time F=1.44, df=3, 160,p=0.23). Income had no meaningful effects when added as a covariate (p<0.0003 for theinteraction).

DiscussionThe current study assessed the effects of neuroticism and acculturation on the CAR inMexican Americans while utilizing a carefully constructed sample monitoring system. Thisis the first study to distinguish the effects of neuroticism from the effects of acculturativestress on the CAR in Mexican Americans, and thus the first to demonstrate that lessacculturated Mexican Americans with greater Mexican orientation who were low inneuroticism demonstrated a magnitude of response and pattern of change over timeindicating a greater CAR compared to their highly acculturated counterparts with highneuroticism. Greater neuroticism was associated with an attenuated CAR in highlyacculturated Mexican Americans with greater Anglo orientation. Particularly dramaticeffects on the CAR were seen in highly acculturated Mexican Americans with highneuroticism. In that group, the CAR virtually disappeared, and changes in cortisol over timewere not significant. Notably, attenuation of the CAR was independent of the effects ofclinical depression and after controlling for childhood trauma, age and gender.

The current study has a number of strengths. Neuroticism increases the risk for depression inadulthood (Duggan et al., 1995; Fanous et al., 2002; Kendler et al., 1993, 2004; Roberts andKendler, 1999) and depression alters HPA axis dynamics including the CAR (Heim andNemeroff, 2001; Huber et al., 2006; Pruessner et al., 2003; Stetler and Miller, 2005).However, the current findings are not attributable to the effects of clinical depression giventhat participants in the study were excluded if they met diagnostic criteria for current orlifetime depression. In addition, we statistically controlled for the potential effects ofchildhood trauma, age, and gender on the CAR, all of which are known to influence theCAR. Participants were excluded for current antidepressant use, alcohol, nicotine and otherdrug use disorders to minimize the effects of these potential confounds on the HPA axis(Aihara et al., 2007; Badrick et al., 2007; Harris et al., 2007; Steptoe and Ussher, 2006;

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Wand and Dobs, 1991). To mitigate the effects of estrogen and progesterone on the HPAaxis female participants were excluded for oral contraceptive use and sampled during theearly follicular phase (Kirschbaum et al., 1999; Kudielka and Kirschbaum, 2005).Measurements were performed with strict reference to time of awakening (Federenko et al.,2004; Williams et al., 2005; Wust et al., 2000b) and sampling error was minimized throughthe use of electronic monitoring devices (Broderick et al., 2004; Kudielka et al., 2003;review in Hansen et al., 2008). Moreover, the current study measured the CAR across twoseparate sampling sessions to minimize the influence of fluctuating situational factors on theCAR, and to maximize trait specificity (Buske-Kirschbaum et al., 2007; Clow et al., 2004;Hellhammer et al., 2007; Kunz-Ebrecht et al., 2004; Scholtz et al., 2004; Thorn et al., 2006).

Our finding that greater neuroticism is associated with an attenuated CAR in highlyacculturated Mexican Americans with greater Anglo orientation is consistent with findingsshowing an attenuated CAR in individuals with high neuroticism (Hauner et al., 2008),severe stress (Thorn et al., 2006) and burnout (Pruessner et al., 1999; Sonnenschein et al.,2007). Findings support previous studies showing the relationship between greateracculturative stress and negative health outcomes may be modulated by personality variablesin various cultural groups (Lazarus and Opton, 1966) including Mexican-Americans(Cervantes & Castro, 1985; Mena, et al., 1987; Miranda & Matheny, 2000; Padilla et al.,1986; Pearlin & Schooler, 1978). However, findings from the current study are in contrastwith studies showing neuroticism is associated with an enhanced CAR. For example, in astudy of 30, healthy males and females, aged 21 to 57, neuroticism was associated with anenhanced CAR after controlling for past and current Axis 1 disorders (depression), medicalillness and medication use, but not for childhood trauma, oral contraceptives, age ordetermination of sampling compliance (Portella et al., 2005). Similar results were reportedin a study of 144 newly diagnosed, female breast cancer patients and controls, aged 30 to 75,in which neuroticism was associated with greater early morning peak cortisol levels, butonly in the breast cancer patients, and with no controls for childhood trauma, age, oralcontraceptive use, menstrual cycle phase, or determination of sampling compliance(Vedhara et al., 2006). Additionally, the effects of traits highly correlated with neuroticism(e.g., negative affect and attachment anxiety) on the CAR were assessed in a study of 334healthy males and females, aged 18 to 54, in which greater negative affect was associatedwith a larger rise in CAR, but only in males, after controlling for depression, medical illness,age, and sampling compliance, but not for childhood trauma, oral contraceptive use,menstrual phase or hormone replacement (Polk et al., 2005). Conflicting results may be dueto failure to control for crucial factors known to influence the CAR (e.g., childhood trauma,age, gender, variations in the measurement of cortisol, determination of samplingcompliance and the potential moderating effects of exposure to chronic stressors), that werecontrolled in the current study.

Current results showing attenuation of the CAR in highly acculturated Mexican Americanswith greater Anglo orientation suggests that acculturation can lead to the dysregulation ofcrucial physiological stress response systems for some Mexican Americans. Consistent withthe attenuation hypothesis, it is plausible that, severe challenges and stressors associatedwith acculturation are associated with the development of allostatic load over time inparticularly vulnerable individuals leading to alterations of the circadian rhythm anddecreased adrenal capacity (Pruessner et al., 1999; Sapolsky, 2004). Similar effects on theaxis are observed in individuals exposed to discrimination (Steptoe et al., 2007) anddownward socioeconomic trajectories (Gustafsson et al., 2009).

There are some limitations to the current study worth noting. The retrospective designprohibits determination of causal pathways between neuroticism, acculturation andattenuation of the CAR. A significant portion of the sample reported a relatively low

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household income; therefore it is possible that family income contributed to a diminishedCAR, possibly due to material hardship (Anderson and Armstead, 1995; Ranjit et al., 2005).However, we believe it is unlikely that our findings are due to the overall lower income inour sample because there was no correlation between parental income and either neuroticism(r=0.08, df=55, p=0.52) or acculturation (r = 0.21, df = 55, p = 0.11) and the effects ofsocioeconomic parameters on cortisol secretion and the CAR indicate an inverse relationship(Bennett et al., 2004; Kapuku et al., 2002; Kunz-Ebrecht et al., 2004; Lupien et al., 2000,2001; Rosmond and Bjorntorp, 2000; Wright and Steptoe, 2005). However, it is possiblethat the burden imposed by increased economic burden contributed to an initialenhancement transitioning over time to attenuation of the CAR.

Determination of the precise developmental mechanisms underlying the relationshipbetween neuroticism and attenuation of the CAR in highly acculturated Mexican Americansis beyond the scope of the current study. However, it is possible that individuals with highlevels of neuroticism are particularly susceptible to increased neuroendocrine reactivityfollowing exposure to the chronic stressors associated with acculturation leading tosignificant attenuation of the CAR. Consistent with the attenuation hypothesis, exposure tochronic stress may lead to enhanced stress reactivity, resulting in down regulation of centralglucocorticoid receptors over time. The hypothalamic CRF system may undergodesensitization transitioning from a system with increased stress responsivity to a systemwith attenuated cortisol responses to stress and an attenuated CAR (Gunnar and Vazquez,2001; Heim et al., 2008; Susman, 2006; Trickett et al., 2010). Alternatively, it is alsoplausible that exposure to stressors associated with acculturation in Mexican Americans withhigh levels of neuroticism produces a blunted CAR with no antecedent HPA axishyperactivity.

Taken together, these findings show that less acculturated Mexican Americans with greaterMexican orientation who were low in neuroticism demonstrated a greater CAR compared totheir more acculturated counterparts with higher neuroticism. In highly acculturatedMexican Americans with greater Anglo orientation and high neuroticism, the CAR wassignificantly attenuated.

RESEARCH HIGHLIGHTS

• We examined the effects of neuroticism, and acculturation on cortisolawakening response (CAR).

• 59 healthy Mexican American adults, ages 18 to 38 years, enrolled in the study.

• Salivary samples were collected at awakening, and 30, 45, and 60 minutesthereafter on 2 days.

• Results showed a significant Neuroticism × Acculturation × Time interaction.

• The CAR was attenuated in highly acculturated Mexican Americans with highneuroticism.

AcknowledgmentsThis work was supported by NIH grant R24-MH070636-01A1 (DLM).

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Wust S, Federenko I, Hellhammer DH, Kirschbaum C. Genetic factors, perceived chronic stress, andthe free cortisol response to awakening. Psychoneuroendocrinology. 2000a; 25:707–720.[PubMed: 10938450]

Wust S, Wolf J, Hellhammer DH, Federenko I, Schommer N, Kirschbaum C. The cortisol awakeningresponse - normal values and confounds. Noise Health. 2000b; 2:79–88. [PubMed: 12689474]

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Figure 1. The effects of the interaction of Neuroticism (N) by Acculturation (A) by Time on theCortisol Awakening Response (CAR) in Mexican AmericansThe effects of the Time × Neuroticism × Acculturation interaction on the CortisolAwakening Response (CAR). The three-factor Neuroticism by Acculturation by Timeinteraction was significant (p = 0.0026) when the Neuroticism and Acculturation scales wereincluded as dimensional covariates. To display the result, Neuroticism and Acculturationscales were split at their means to create dichotomous indicators (low, high), and the datareanalyzed using these classifications in a 2 (Neuroticism) × 2 (Acculturation) × 4 (Time)mixed effects regression model. Participants who were low on both dimensions had a greaterCAR (a test of change over time in this group yielded (F = 14.75, df = 3,162, p<0.0001). Incontrast, greater neuroticism was associated with an attenuated CAR in highly acculturatedMexican Americans with greater Anglo orientation (test for change over time yielded (F=1.07, df = 3, 162, p = 0.36). The graphic displays the exponentiated means of the log-transformed data (antilogs are presented to put the data in its original scale).

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Table 1Sample demographics for Mexican American adult, males and females

Table 1 presents demographic characteristics of the sample. Participants were single, bilingual, healthy, youngadult male and female college students, with an even distribution among first, second and third generationalgroups.

OverallSample Females Males

Gender (n=59) 33 (56%) 26 (44%)

Age (Median and Range) 20 (18–38) 21(18–38) 19 (18–37)

Body Mass Index (Mean and SD) 24.39 ± 4.19 24.54 ± 4.5 24.20 ± 3.8

Parental/Participant Income

Below 40K 33 (55.9%) 22 (66.7%) 11 (42.3%)

40K–80K 16 (27.1%) 7 (21.2%) 9 (34.6%)

80K and above 10(17.0%) 4 (12.1%) 6 (23.1%)

Parental Occupational Status

Executive 10 (16.9%) 5(15.2%) 5 (19.2%)

Administrative/Management 18 (30.5%) 11 (33.3%) 7 (27.0%)

Clerical/Skilled Manual Labor 20 (34.0%) 11 (33.3%) 9 (34.6%)

Semi-skilled or Unskilled Labor 11 (18.6%) 6 (18.2%) 5 (19.2%)

Participant Education Level

≤12 years 4 (6.8%) 2 (6.1%) 2(7.7%)

13 years 23 (39.0%) 11 (33.3%) 12 (46.1%)

14 years 13 (22.0%) 11 (33.3%) 2 (7.7%)

15 years 6 (10.2%) 2 (6.1%) 4 (15.4%)

≥16 year 13 (22.0%) 7 (21.2%) 6 (23.1%)

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Table 2Neuroticism factor and facet scores for Mexican American adults (NEO-PI-R; Means andSD)

Table 2 reports Neuroticism factor and facet scores for the sample of Mexican American adults (NEO-PI-R;Costa and McCrae, 1992). Overall raw scores for the current sample were consistent with norms published forpredominately Caucasian college samples (Costa and McCrae, 1992a) and with scores previously reported fora Mexican American, college sample (Mangold et al., 2007). There were no significant differences on theneuroticism factor and facet scores among males and females

N=59 College Aged Males College Aged Females

Factors

Neuroticism (N) 92.24 (26.5) 86.9 (23.3)

Facets

Anxiety (N1) 14.8 (5.3) 16.9 (5.2)

Angry Hostility (N2) 15.4 (5.2) 13.7 (5.5)

Depression (N3) 14.8 (6.3) 13.5 (6.0)

Self-Consciousness (N4) 16.6 (5.3) 14.6 (5.2)

Impulsiveness (N5) 19.0 (4.3) 16.2 (4.3)

Vulnerability (N6) 13.2 (5.5) 12.1 (4.1)

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Table 3The effects of neuroticism and acculturation on the Cortisol Awakening Response (CAR)in Mexican Americans

Table 3 presents results from the mixed effects regression model examining the effects of neuroticism andacculturation on the Cortisol Awakening Response (CAR) in Mexican Americans. Results showed asignificant Neuroticism × Acculturation × Time interaction (p = .0026). Mexican Americans with greaterMexican orientation and low neuroticism demonstrated a healthy, robust CAR, while the CAR wassignificantly attenuated in Mexican Americans with relatively greater Anglo orientation and high neuroticism.

Effect df F P

Neuroticism (N) 1,59 2.01 0.16

Acculturation (A) 1,59 3.00 0.09

N × A 1,59 0.09 0.76

Time 3,163 28.02 < 0.0001

N × Time 3,163 1.18 0.32

A × Time 3,163 5.42 0.0014

N × A × Time 3,163 4.93 0.0026

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